ABSTRACT
The development of a technique for laser measurement of fPhotosystem II (PS II) photochemical characteristics of phytoplankton and terrestrial vegetation from an airborne platform is described. Results of theoretical analysis and experimental study of pump-and-probe measurement of the PS II functional absorption cross-section and photochemical quantum yield are presented. The use of 10 ns probe pulses of PS II sub-saturating intensity provides a significant, up to 150-fold, increase in the fluorescence signal compared to conventional 'weak-probe' protocol. Little effect on the fluorescence yield from the probe-induced closure of PS II reaction centers is expected over the short pulse duration, and thus a relatively intense probe pulse can be used. On the other hand, a correction must be made for the probe-induced carotenoid triplet quenching and singlet-singlet annihilation. A Stern-Volmer model developed for this correction assumes a linear dependence of the quenching rate on the laser pulse fluence, which was experimentally validated. The PS II saturating pump pulse fluence (532 nm excitation) was found to be 10 and 40 mumol quanta m(-2) for phytoplankton samples and leaves of higher plants, respectively. Thirty mus was determined as the optimal delay in the pump-probe pair. Our results indicate that the short-pulse pump-and-probe measurement of PS II photochemical characteristics can be implemented from an airborne platform using existing laser and LIDAR technologies.
ABSTRACT
Initial results of the airborne LIDAR measurement of photochemical quantum yield, Phi(Po), and functional absorption cross-section, sigma(PS II), of Photosystem II (PS II) are reported. NASA's AOL3 LIDAR was modified to implement short-pulse pump-and-probe (SP-P&P) LIDAR measurement protocol. The prototype system is capable of measuring a pump-induced increase in probe-stimulated chlorophyll fluorescence, DeltaF/F(sat), along with the acquisition of ;conventional' LIDAR-fluorosensor products from an operational altitude of 150 m. The use of a PS II sub-saturating probe pulse increases the response signal but also results in excessive energy quenching (EEQ) affecting the DeltaF/F(sat) magnitude. The airborne data indicated up to a 3-fold EEQ-caused decline in DeltaF/F(sat), and 2-fold variability in the EEQ rate constant over a spatial scale a few hundred kilometers. Therefore, continuous monitoring of EEQ parameters must be incorporated in the operational SP-P&P protocol to provide data correction for the EEQ effect. Simultaneous airborne LIDAR measurements of Phi(Po) and sigma(PS II) with EEQ correction were shown to be feasible and optimal laser excitation parameters were determined. Strong daytime DeltaF/F(sat) decline under ambient light was found in the near-surface water layer over large aquatic areas. An example of SP-P&P LIDAR measurement of phytoplankton photochemical and fluorescent characteristics in the Chesapeake Bay mouth is presented. Prospects for future SP-P&P development and related problems are discussed.
ABSTRACT
BACKGROUND: Active fluorescence techniques are becoming commonly used to monitor the state of the photosynthetic apparatus in natural populations of phytoplankton, but at present these are bulk water measurements that average all the fluorescent material in each sample. Here we describe two instruments that combine individual-cell "pump-during-probe" (PDP) measurements of chlorophyll (Chl) fluorescence induction, on the time scale of 30 to 100 micros, with flow cytometric or visual characterization of each cell. METHODS: In the PDP flow cytometer, we measure Chl fluorescence yield as a function of time during a 150 micros excitation flash provided by an argon ion laser; each particle is subsequently classified as in a conventional flow cytometer. In the PDP microfluorometer, individual cells in a sample chamber are visually identified, and fluorescence excitation is provided by a blue light-emitting diode that can be configured to provide a saturating flash and also a subsequent series of short flashlets. This sequence allows both saturation and relaxation kinetics to be monitored. RESULTS: Phytoplankton from natural samples and on-deck iron-enrichment incubation experiments in the Southern Ocean were examined with each PDP instrument, providing estimates of the potential quantum yield of photochemistry and the functional absorption cross section for photosystem 2, for either individuals (for cells larger than a few micrometers) or populations (for smaller cells). CONCLUSIONS: Results from initial field applications indicate that single-cell PDP measurements can be a powerful tool for investigating the nutritional state of phytoplankton cells and the regulation of phytoplankton growth in the sea.
